高通量优化主要编辑介导内源蛋白标记,用于聚合蛋白质定位成像的聚合成像
Henry M Sanchez1,2,3, Tomer Lapidot1,2, Ophir Shalem1,2
1Center for Cellular and Molecular Therapeutics, Children's Hospital of Philadelphia, Philadelphia, PA 19104, USA.
bioRxiv : the preprint server for biology
|September 30, 2024
概括
我们开发了一种可扩展的方法,使用主要编辑来标记细胞内的蛋白质,从而能够大规模测量它们的位置. 这项技术允许在各种细胞条件下分析蛋白质组的组织和动态.
科学领域:
- 细胞生物学 细胞生物学
- 分子生物学分子生物学
- 基因组学就是基因组学.
背景情况:
- 细胞下蛋白质组织对于理解细胞状态和基因功能至关重要.
- 目前的技术缺乏在规模上准确测量亚细胞蛋白位址的能力.
研究的目的:
- 开发一种可扩展的技术,用于测量亚细胞蛋白位址.
- 为了实现蛋白质组组织的高通量分析.
主要方法:
- 开发了一种使用主要编辑的聚合内源蛋白标记方法.
- 创建了17,280个主要编辑指导RNA (pegRNAs) 的聚合库,针对60种内源蛋白质.
- 集成的 in-situ pegRNA 测序与深度学习图像分析.
主要成果:
- 确定了影响标记效率的基因组和pegRNA特征,例如表观遗传状态和转录相互作用.
- 开发了一个计算模型来预测 pegRNA 设计,以实现高效的突入.
- 在单次转导后,证明了对众多蛋白质细胞下蛋白质局部化的并行探索.
结论:
- 开发的基于主要编辑的方法可以进行可扩展的,高吞吐量测量亚细胞蛋白质定位.
- 这项技术有助于对不同细胞类型和环境条件的蛋白质组动态进行大规模研究.
- 为未来的蛋白质组组织和功能研究奠定了基础.
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